Fig 6
![Early cell death of zebrin II Purkinje cells in Herc2 +/530 mutants. Microphotographs of immunohistochemical staining with antibodies against HERC2, calbindin (CaBP) and aldolase C/zebrin II in coronal sections through the vermis (A-B), and paravermis (C-E) [see 99, 112] of the 9-month-old HERC2 +/530 cerebellum. Purkinje cells express HERC2 protein (A) and cell death follows a parasagittal gradient in which the number of dead Purkinje cells increases from the vermis to the paravermal zones (compare asterisks A-C). Signs of Purkinje cell damage like axonal torpedoes (D, arrows) and thick dendritic debris (D-E, arrowheads) were consistently found. The arrows in B indicate calbindin (CaBP) and zebrin II double labeled Purkinje cells, while arrowheads show single CaBP expressing Purkinje cells. Bars = 1000 µm (A-C) and 50 µm (D-E).](profile/Sara_Bachiller/publication/356591317/figure/fig6/AS:11431281176957923@1690301291918/Early-cell-death-of-zebrin-II-Purkinje-cells-in-Herc2-530-mutants-Microphotographs-of.png)
Early cell death of zebrin II Purkinje cells in Herc2 +/530 mutants. Microphotographs of immunohistochemical staining with antibodies against HERC2, calbindin (CaBP) and aldolase C/zebrin II in coronal sections through the vermis (A-B), and paravermis (C-E) [see 99, 112] of the 9-month-old HERC2 +/530 cerebellum. Purkinje cells express HERC2 protein (A) and cell death follows a parasagittal gradient in which the number of dead Purkinje cells increases from the vermis to the paravermal zones (compare asterisks A-C). Signs of Purkinje cell damage like axonal torpedoes (D, arrows) and thick dendritic debris (D-E, arrowheads) were consistently found. The arrows in B indicate calbindin (CaBP) and zebrin II double labeled Purkinje cells, while arrowheads show single CaBP expressing Purkinje cells. Bars = 1000 µm (A-C) and 50 µm (D-E).
Source publication
The HERC protein family is one of three subfamilies of Homologous to E6AP C-terminus (HECT) E3 ubiquitin ligases. Six HERC genes have been described in humans, two of which encode Large HERC proteins -HERC1 and HERC2- with molecular weights above 520 kDa that are constitutively expressed in the brain. There is a large body of evidence that mutation...
Citations
... HERC2 is a ubiquitin ligase involved in various physiological processes, including membrane trafficking, inflammation, immune response, DNA repair, and the cellular stress response [8][9][10][11]. In situ hybridization studies in mice have provided evidence of Herc2 expression in different areas of the nervous system such as the isocortex, hippocampus, thalamus, hypothalamus, midbrain, pons, medulla and cerebellum [12]. The HERC2 gene is located near an imprinting region of chromosome 15 associated with neurodevelopmental disorders like Angelman syndrome. ...
Sequence variants in the HERC2 gene are associated with a significant reduction in HERC2 protein levels and cause a neurodevelopmental disorder known as the HERC2-related disorder, which shares clinical features with Angelman syndrome, including global developmental delay, intellectual disability, autism, and movement disorders. Remarkably, the HERC2 gene is commonly deleted in individuals with Angelman syndrome, suggesting a potential contribution of HERC2 to the pathophysiology of this disease. Given the known critical role of autophagy in brain development and its implication in neurodevelopmental diseases, we undertook different experimental approaches to monitor autophagy in fibroblasts derived from individuals affected by the HERC2-related disorder. Our findings reveal alterations in the levels of the autophagy-related protein LC3. Furthermore, experiments with lysosomal inhibitors provide confirmation of an upregulation of the autophagy pathway in these patient-derived cells. Mechanistically, we corroborate an interaction between HERC2 and the deubiquitylating enzyme USP20; and demonstrate that HERC2 deficiency leads to increased USP20 protein levels. Notably, USP20 upregulation correlates with enhanced stability of the autophagy initiating kinase ULK1, highlighting the role of HERC2 as an autophagy regulator factor through the USP20-ULK1 axis. Moreover, we show that p38 acts as a modulator of this pathway, since p38 activation disrupts HERC2-USP20 interaction, leading to increased USP20 and LC3-II protein levels. Together, these findings uncover a previously unknown role for HERC2 in autophagy regulation and provide insights into the pathomolecular mechanisms underlying the HERC2-related disorder and Angelman syndrome.
... Furthermore, among the different genes that were differentially expressed, we also identified the large Herc family members Herc1 and Herc2. Though their temporal changes in terms of gene expression were milder, when compared to those of UBE3A or Nedd4L, large Herc members have recently gained relevance because when their encoding genes are mutated, they are associated with clinical syndromes closely related to NDDs, resulting in intellectual disability, dementia, epileptic seizures, and/or signs of autism [59]. Murine models have helped to clarify the mechanisms underlying the phenotypes and progressive degeneration of Purkinje cells; defects at the neuromuscular junction and impaired motor control have been described [60]. ...
Human brain development involves a tightly regulated sequence of events that starts shortly after conception and continues up to adolescence. Before birth, neurogenesis occurs, implying an extensive differentiation process, sustained by changes in the gene expression profile alongside proteome remodeling, regulated by the ubiquitin proteasome system (UPS) and autophagy. The latter processes rely on the selective tagging with ubiquitin of the proteins that must be disposed of. E3 ubiquitin ligases accomplish the selective recognition of the target proteins. At the late stage of neurogenesis, the brain starts to take shape, and neurons migrate to their designated locations. After birth, neuronal myelination occurs, and, in parallel, neurons form connections among each other throughout the synaptogenesis process. Due to the malfunctioning of UPS components, aberrant brain development at the very early stages leads to neurodevelopmental disorders. Through deep data mining and analysis and by taking advantage of machine learning-based models, we mapped the transcriptomic profile of the genes encoding HECT- and ring-between-ring (RBR)-E3 ubiquitin ligases as well as E2 ubiquitin-conjugating and E1 ubiquitin-activating enzymes during human brain development, from early post-conception to adulthood. The inquiry outcomes unveiled some implications for neurodevelopment-related disorders.
... The physiological relevance of the Large HERCs is illustrated by their involvement in different diseases as shown in Figure 1. Although expressed in different tissues, Large HERCs are notably expressed throughout different areas of the nervous system [17]. Therefore, it is not surprising that most of the diseases with which these ubiquitin ligases are related are of a neurological nature. ...
... HERC1: several cancers [18]; Macrocephaly, Dysmorphic Facies and PsychoMotor Retardation (MDFPMR) syndrome [19][20][21][22][23]; autism spectrum disorder [24,25]; Parkinson's disease [26]; schizophrenia [27]; febrile seizures [28]; neuropathic periphery disease [29]; COVID-19 combined with major depression disorder (COVID-19-MDD) [30]; acquired immunodeficiency syndrome (AIDS) [31]; diabetes [32]; cardiovascular disease [33]; osteopenia [34]. HERC2: several cancers [18]; HERC2 Angelman-like syndrome [35][36][37][38][39][40][41][42][43][44]; autism spectrum disorder [17]; Parkinson's disease [45]; agenesis of the corpus callosum (ACC) [46]; brain arteriovenous malformation (BAVM) [47]; diabetic cerebral ischemia-reperfusion (I/R) injury [48]; central precocious puberty [49]; refractive astigmatism [50]; inflammatory diseases [51][52][53][54][55]; asthma [56]; hypertension [57,58]; skin conditions [59][60][61]. ...
... Whether the alterations in these MAPK signaling pathways are associated with clinical outcomes in the neurodevelopmental disorders caused by mutations in HERC1 or HERC2 still requires further research. Even so, dysfunctions of the MAPK signaling pathways have already been implicated in several neurodevelopmental disorders, for instance autism spectrum disorder [96,97], which is also manifested in the syndromes caused by Large HERC mutations [17]. In addition, MAPK pathways are also associated with neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease and amyotrophic lateral sclerosis [98,99]. ...
Protein ubiquitylation acts as a complex cell signaling mechanism since the formation of different mono- and polyubiquitin chains determines the substrate’s fate in the cell. E3 ligases define the specificity of this reaction by catalyzing the attachment of ubiquitin to the substrate protein. Thus, they represent an important regulatory component of this process. Large HERC ubiquitin ligases belong to the HECT E3 protein family and comprise HERC1 and HERC2 proteins. The physiological relevance of the Large HERCs is illustrated by their involvement in different pathologies, with a notable implication in cancer and neurological diseases. Understanding how cell signaling is altered in these different pathologies is important for uncovering novel therapeutic targets. To this end, this review summarizes the recent advances in how the Large HERCs regulate the MAPK signaling pathways. In addition, we emphasize the potential therapeutic strategies that could be followed to ameliorate the alterations in MAPK signaling caused by Large HERC deficiencies, focusing on the use of specific inhibitors and proteolysis-targeting chimeras.
... HERC2 has been widely reported as an E3 ubiquitin ligase [26,27]. We thus first evaluated whether HERC2 participated in proteasome-mediated PTP1B degradation. ...
... The E3 ligases have been classified into three families, the HECT-type, RING-type, and U-box-type E3 ligases [57]. HERC2 belongs to the HECT-type E3 ubiquitin family that mediated protein degradation [26,27]. The RING-type ubiquitin ligase tripartite motif 18 (TRIM18) and U-box-type ubiquitin ligase precursor RNA processing-19β (PRP19β) have been reported to regulate PTP1B levels by ubiquitindependent proteasomal degradation under STAT3 activation [58,59]. ...
Background:
Hepatic inflammation is a common initiator of liver diseases and considered as the primary driver of hepatocellular carcinoma (HCC). However, the precise mechanism of inflammation-induced HCC development and immune evasion remains elusive and requires extensive investigation. This study sought to identify the new target that is involved in inflammation-related liver tumorigenesis.
Methods:
RNA-sequencing (RNA-seq) analysis was performed to identify the differential gene expression signature in primary human hepatocytes treated with or without inflammatory stimulus. A giant E3 ubiquitin protein ligase, HECT domain and RCC1-like domain 2 (HERC2), was identified in the analysis. Prognostic performance in the TCGA validation dataset was illustrated by Kaplan-Meier plot. The functional role of HERC2 in HCC progression was determined by knocking out and over-expressing HERC2 in various HCC cells. The precise molecular mechanism and signaling pathway networks associated with HERC2 in HCC stemness and immune evasion were determined by quantitative real-time PCR, immunofluorescence, western blot, and transcriptomic profiling analyses. To investigate the role of HERC2 in the etiology of HCC in vivo, we applied the chemical carcinogen diethylnitrosamine (DEN) to hepatocyte-specific HERC2-knockout mice. Additionally, the orthotopic transplantation mouse model of HCC was established to determine the effect of HERC2 during HCC development.
Results:
We found that increased HERC2 expression was correlated with poor prognosis in HCC patients. HERC2 enhanced the stemness and PD-L1-mediated immune evasion of HCC cells, which is associated with the activation of signal transducer and activator of transcription 3 (STAT3) pathway during the inflammation-cancer transition. Mechanically, HERC2 coupled with the endoplasmic reticulum (ER)-resident protein tyrosine phosphatase 1B (PTP1B) and limited PTP1B translocation from ER to ER-plasma membrane junction, which ameliorated the inhibitory role of PTP1B in Janus kinase 2 (JAK2) phosphorylation. Furthermore, HERC2 knockout in hepatocytes limited hepatic PD-L1 expression and ameliorated HCC progression in DEN-induced mouse liver carcinogenesis. In contrast, HERC2 overexpression promoted tumor development and progression in the orthotopic transplantation HCC model.
Conclusion:
Our data identified HERC2 functions as a previously unknown modulator of the JAK2/STAT3 pathway, thereby promoting inflammation-induced stemness and immune evasion in HCC.
... Of the six HERC genes described in humans, two (HERC1 and HERC2) encode large HERC proteins with molecular weights above 520 kDa. Of these, HERC1 somatic mutations have been identified in cancers such as leukemia, breast cancer, and non-melanoma skin cancer [9,10] and germline mutations have been reported in neuronal disorders [11,12]. In this context, genetic studies have related HERC1 mutations with autism spectrum disorders, considering it a predictor of autism risk [13,14]. ...
... In this context, genetic studies have related HERC1 mutations with autism spectrum disorders, considering it a predictor of autism risk [13,14]. The common features observed in individuals with homozygous or compound heterozygous mutations in the HERC1 gene led to the identification of the autosomal recessive neurodevelopmental disorder MDFPMR syndrome (macrocephaly, dysmorphic facies, and psychomotor retardation) (OMIM # 617011) [15][16][17][18][19]. Altered bone homeostasis could account for several features observed in these individuals, such as macrocephaly, dysmorphic facies, prominent forehead, long fingers, and vertebral column abnormalities [11]. Most of the HERC1 mutations associated with MDFPMR syndrome are frameshift mutations that cause truncated HERC1 proteins and a loss of function [15][16][17][18]. ...
Bone remodeling is a continuous process between bone-forming osteoblasts and bone-resorbing osteoclasts, with any imbalance resulting in metabolic bone disease, including osteopenia. The HERC1 gene encodes an E3 ubiquitin ligase that affects cellular processes by regulating the ubiquitination of target proteins, such as C-RAF. Of interest, an association exists between biallelic pathogenic sequence variants in the HERC1 gene and the neurodevelopmental disorder MDFPMR syndrome (macrocephaly, dysmorphic facies, and psychomotor retardation). Most pathogenic variants cause loss of HERC1 function, and the affected individuals present with features related to altered bone homeostasis. Herc1-knockout mice offer an excellent model in which to study the role of HERC1 in bone remodeling and to understand its role in disease. In this study, we show that HERC1 regulates osteoblastogenesis and osteoclastogenesis, proving that its depletion increases gene expression of osteoblastic makers during the osteogenic differentiation of mesenchymal stem cells. During this process, HERC1 deficiency increases the levels of C-RAF and of phosphorylated ERK and p38. The Herc1-knockout adult mice developed imbalanced bone homeostasis that presented as osteopenia in both sexes of the adult mice. By contrast, only young female knockout mice had osteopenia and increased number of osteoclasts, with the changes associated with reductions in testosterone and dihydrotestosterone levels. Finally, osteocytes isolated from knockout mice showed a higher expression of osteocytic genes and an increase in the Rankl/Opg ratio, indicating a relevant cell-autonomous role of HERC1 when regulating the transcriptional program of bone formation. Overall, these findings present HERC1 as a modulator of bone homeostasis and highlight potential therapeutic targets for individuals affected by pathological HERC1 variants.
... HERC1, a ubiquitin ligase, contributes to autophagy/lysosome and proteasome pathways that lead to protein degradation [36][37][38] . In the peripheral nervous system, HERC1 mutations alter presynaptic membrane dynamics, resulting in delayed neurotransmission and impaired movement and learning 39 . According to GO enrichment analysis, the most common genes were involved in autophagy-related endomembrane systems such as lysosomes and endoplasmic reticulum membranes and neurological signaling. ...
Objective:
A lack of objective biomarkers is preventing the screening and diagnosis of COVID-19 combined with major depression disorder (COVID-19-MDD). The purpose of this study was to identify diagnostic biomarkers and gene regulatory mechanisms associated with autophagy; a crucial process significantly involved in the pathogenesis of COVID-19-MDD.
Materials and methods:
In this study, differentially expressed genes (DEGs) were screened using GSE98793 from the GEO2R analysis (GEO) database, and intersected with the COVID-19-related gene (CRGs) and autophagy-related genes (ARGs) to obtain common genes involved in. Then, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses of these common genes were performed. Subsequently, the transcription factor (TF)-gene regulatory network and comorbidity network were constructed. In addition, 10 drug candidates were screened using the DSigDB database. To identify diagnostic markers, we used LASSO regression.
Results:
In total, 13 common genes were screened, which were primarily enriched in lysosomes, endoplasmic reticulum membranes, and other endomembrane systems also associated with autophagy. Additionally, these genes were involved in neurological cell signaling and have a functional role in pathways related to vascular endothelial growth factor, tyrosine kinase, autophagy, inflammation, immunity, and carcinogenesis. Tumors and psychiatric disorders were the most highly linked diseases to COVID-19. Finally, ten drug candidates and eight diagnostic markers (STX17, NRG1, RRAGD, XPO1, HERC1, HSP90AB1, EPHB2, and S1PR3) were screened.
Conclusions:
This is the first study to screen eight diagnostic markers and construct a gene regulatory network for COVID-19-MDD from the perspective of autophagy. The findings of our study provide novel insights into the diagnosis and treatment of COVID-19-MDD.
... Besides, HERC2 is essential during embryonic development and plays an important role in regulating motor coordination [13]. Moreover, it is highly expressed in the nervous system and has been linked with hereditary neurodegenerative disorders [14]. Biallelic HERC2 variants associated with HERC2 Angelman-like syndrome include missense and frameshift mutations with a premature stop codon that result in a loss of function. ...
... Several recessive mutations affecting the HERC2 gene cause developmental delay with Angelman-like features [14,19]. Knowing how pathologic HERC2 variants affect intracellular signalling could reveal the underlying pathology and identify possible therapies. ...
HERC2 gene encodes an E3 ubiquitin ligase involved in several cellular processes by regulating the ubiquitylation of different protein substrates. Biallelic pathogenic sequence variants in the HERC2 gene are associated with HERC2 Angelman-like syndrome. In pathogenic HERC2 variants, complete absence or marked reduction in HERC2 protein levels are observed. The most common pathological variant, c.1781C > T (p.Pro594Leu), encodes an unstable HERC2 protein. A better understanding of how pathologic HERC2 variants affect intracellular signalling may aid definition of potential new therapies for these disorders. For this purpose, we studied patient-derived cells with the HERC2 Pro594Leu variant. We observed alteration of mitogen-activated protein kinase signalling pathways, reflected by increased levels of C-RAF protein and p38 phosphorylation. HERC2 knockdown experiments reproduced the same effects in other human and mouse cells. Moreover, we demonstrated that HERC2 and RAF proteins form molecular complexes, pull-down and proteomic experiments showed that HERC2 regulates C-RAF ubiquitylation and we found out that the p38 activation due to HERC2 depletion occurs in a RAF/MKK3-dependent manner. The displayed cellular response was that patient-derived and other human cells with HERC2 deficiency showed higher resistance to oxidative stress with an increase in the master regulator of the antioxidant response NRF2 and its target genes. This resistance was independent of p53 and abolished by RAF or p38 inhibitors. Altogether, these findings identify the activation of C-RAF/MKK3/p38 signalling pathway in HERC2 Angelman-like syndrome and highlight the inhibition of RAF activity as a potential therapeutic option for individuals affected with these rare diseases.
... For instance, it is well known that mutations in HERC2 produce clinical syndromes in which key neurodevelopmental events are altered, resulting in intellectual disability and other neurological disorders [34]. Genetic variation in PDE1C ...
Objective:
Bipolar Disorder (BD) is an heritable chronic mental disorder causing psychosocial impairment, affecting patients with depressive/manic episodes. The familial transmission of BD does not follow any of the simple Mendelian patterns of inheritance. The aim of this study is to describe a new large family with twelve affected BD members: WES was performed in eight of them, three of which were diagnosed for BD, and one was reported as a "borderline" individual.
Material and methods:
WES data allowed us to select variants in common between the affected subjects, once including and once excluding a "borderline" subject with moderate anxiety and traits of obsessive-compulsive disorder.
Results:
Results were in favor of new predisposing BD genes, electing a heterozygous missense variant in CLN6 resulting in a "borderline" phenotype that if combined with a heterozygous missense variant in ZNF92 is responsible for the more severe BD phenotype. Both rare missense changes are predicted to disrupt the protein function.
Conclusions:
Loss of both alleles in CLN6 causes Neuronal Ceroid Lipofuscinosis, a severe progressive neurological disorder of childhood. Our results indicate that heterozygous CLN6 carriers, previously reported as healthy, may be susceptible to bipolar disorder late in life if associated with additional variants in ZNF92.
Lysosomes are central to metabolic homeostasis. The microphthalmia bHLH-LZ transcription factors (MiT/TFEs) family members MITF, TFEB, and TFE3 promote the transcription of lysosomal and autophagic genes and are often deregulated in cancer. Here, we show that the GATOR2 complex, an activator of the metabolic regulator TORC1, maintains lysosomal function by protecting MiT/TFEs from proteasomal degradation independent of TORC1, GATOR1, and the RAG GTPase. We determine that in GATOR2 knockout HeLa cells, members of the MiT/TFEs family are ubiquitylated by a trio of E3 ligases and are degraded, resulting in lysosome dysfunction. Additionally, we demonstrate that GATOR2 protects MiT/TFE proteins in pancreatic ductal adenocarcinoma and Xp11 translocation renal cell carcinoma, two cancers that are driven by MiT/TFE hyperactivation. In summary, we find that the GATOR2 complex has independent roles in TORC1 regulation and MiT/TFE protein protection and thus is central to coordinating cellular metabolism with control of the lysosomal-autophagic system.
Lead (Pb) exposure causes immeasurable damage to multiple human systems, particularly the central nervous system (CNS). In this study, human induced pluripotent stem cells (hiPSCs) were differentiated into neural progenitor cells (NPCs) to investigate the neurotoxic effects of Pb. The hiPSCs were treated with 0, 0.5, 1.0, 2.5, 5.0 and 10.0 μmol/L Pb for 7 days, whereas embryoid bodies (EBs) and NPCs were treated with 0, 0.1, 0.5, and 1.0 μmol/L Pb for 7 days. Pb exposure disrupted the cell cycle and caused apoptosis in hiPSCs, EBs, and NPCs. Besides, Pb inhibited the differentiation of NPCs and EBs. Whole exome sequencing revealed 2509, 2413, and 1984 single nucleotide variants (SNVs) caused by Pb in hiPSCs, EBs, and NPCs, respectively. The common mutation sites in the exon region were mostly nonsynonymous mutations. We identified 18, 19, and 18 common deleterious mutations in hiPSCs, EBs, and NPCs, respectively. Additionally, Online Mendelian Inheritance in Man database analysis revealed 30, 20, and 13 genes related to CNS disorders in hiPSCs, EBs, and NPCs, respectively. Our findings suggest that this in vitro model may supplement animal models and be applied to the study of neurodevelopmental toxicity in the future.
